Process for producing pyrrolidine derivative and salt thereof

ABSTRACT

A process for producing a compound of the general formula: ##STR1## wherein R 1  is heterocyclic(lower)alkyl, R 2  is acyl, and 
     R 3  is carboxy(lower)alkyl or protected carboxy(lower)alkyl, 
     which is useful as a medicament or a salt thereof.

This is a Division of application Ser. No. 08/360,679 filed on Dec. 30,1994, now U.S. Pat. No. 5,543,525 which is the U.S. National Stage ofInternational Application No. PCT/JP93/00886 filed Jun. 29, 1993 nowWO94/01400.

TECHNICAL FIELD

The present invention relates to an industrial process for producingpyrrolidine derivative or a salt thereof represented by the followinggeneral formula (I): ##STR2## wherein R¹ is heterocyclic(lower)alkyl, R²is acyl, and

R³ is carboxy(lower)alkyl or protected carboxy(lower) alkyl, and isapplicable in the medical field.

BACKGROUND ART

Pyrrolidine derivatives of the above general formula (I), or saltsthereof and a method for producing them are known as disclosed inJapanese Patent Publication No. 2-152960.

DISCLOSURE OF INVENTION

The method described in the above-mentioned bulletin for the productionof compounds of general formula (I) is disadvantageous in that itrequires a long series of steps complicating the production and providesonly a very low overall yield rendering the production cost high.

The object of this invention is to provide an industrially excellentproduction method for producing compounds of general formula (I) andsalts thereof which is superior to the above known production process insimplicity and in yield.

The method for producing said pyrrolidine derivative (I) according tothis invention is as follows. ##STR3## wherein R¹, R² and R³ are each asdefined above, R⁴ is aryl, and

Y⁻ is an anion.

The starting compound (II) is novel and can be produced by the followingprocesses. ##STR4## wherein R¹ and R² are each as defined above,

R⁵ is carboxy or protected carboxy,

R_(a) ⁵ is protected carboxy,

R⁶ is lower alkyl,

M is an alkali metal,

X¹ and X² are each a leaving group, and

X³ is halogen.

In the above and subsequent descriptions of the present specification,suitable examples and illustrations of the various definitions which thepresent invention include within the scope thereof are explained indetail as follows.

The term "lower" is intended to mean, unless otherwise indicated, 1 to 6carbon atoms.

Suitable "lower alkyl" and "lower alkyl moiety" in the terms"heterocyclic(lower)alkyl", "carboxy(lower)alkyl" and "protectedcarboxy(lower)alkyl" may include straight or branched one having 1 to 6carbon atom(s) such as methyl, ethyl, propyl, isopropyl, butyl,isobutyl, sec-butyl, tert-butyl, pentyl, tert-pentyl, hexyl and thelike, preferably one having 1 to 4 carbon atom(s).

Suitable "aryl" may include phenyl, naphthyl, and the like.

Suitable "heterocyclic moiety" in the term "heterocyclic(lower)alkyl"means saturated or unsaturated hetero-monocyclic or -polycyclic groupcontaining at least one hetero-atom such as oxygen, sulfur and nitrogenatoms. The particularly preferred heterocyclic group may be

unsaturated 3- to 8-membered (more preferably 5- or 6-membered)heteromonocyclic group containing 1 to 4 nitrogen atom(s), such aspyrrolyl, pyrrolinyl, imidazolyl, pyrazolyl, pyridyl and its N-oxide,pyrimidyl, pyrazinyl, pyridazinyl, triazolyl (e.g. 4H-1,2,4-triazolyl,1H-1,2,3-triazolyl, 2H-1,2,3-triazolyl, etc.), tetrazolyl (e.g.1H-tetrazolyl, 2H-tetrazolyl, etc.), dihydrotriazinyl (e.g.4,5-dihydro-1,2,4-triazinyl, 2,5-dihydro-1,2,4-triazinyl, etc.), etc.;

saturated 3- to 8-membered (more preferably 5- or 6-membered)heteromonocyclic group containing 1 to 4 nitrogen atom(s), such aspyrrolidinyl, imidazolidinyl, piperidino, piperazinyl, etc.;

unsaturated condensed heterocyclic group containing 1 to 5 nitrogenatom(s), such as indolyl, isoindolyl, indolizinyl, benzimidazolyl,quinolyl, isoquinolyl, indazolyl, benzotriazolyl, tetrazolopyridyl,tetrazolopyridazinyl (e.g. tetrazolo 1,5-b!pyridazinyl, etc.),dihydrotriazolopyridazinyl, etc.;

unsaturated 3- to 8-membered (more preferably 5- or 6-membered)heteromonocyclic group containing 1 or 2 oxygen atom(s) and 1 to 3nitrogen atom(s), such as oxazolyl, isoxazolyl, oxadiazolyl (e.g.1,2,4-oxadiazolyl, 1,3,4-oxadiazolyl, 1,2,5-oxadiazolyl, etc.), etc.;

saturated 3- to 8-membered (more preferably 5- or 6-membered)heteromonocyclic group containing 1 or 2 oxygen atom(s) and 1 to 3nitrogen atom(s), such as morpholinyl, etc.;

unsaturated condensed heterocyclic group containing 1 or 2 oxygenatom(s) and 1 to 3 nitrogen atom(s), such as benzoxazolyl,benzoxadiazolyl, etc.;

unsaturated 3- to 8-membered (more preferably 5- or 6-membered)hetaromonocyclic group containing 1 or 2 sulfur atom(s) and 1 to 3nitrogen atom(s), such as thiazolyl (e.g. 1,2-thiazolyl, etc.),thiazolinyl, thiadiazolyl (e.g. 1,2,4-thiadiazoyl, 1,3,4-thiadiazolyl,1,2,5-thiadiazolyl, 1,2,3-thiadiazolyl, etc.), etc.;

saturated 3- to 8-membered (more preferably 5- or 6-membered)heteromonocyclic group containing 1 to 2 sulfur atom(s) and 1 to 3nitrogen atom(s), such as thiazolidinyl, etc.;

unsaturated 3- to 8-membered (more preferably 5- or 6-membered)heteromonocyclic group containing one sulfur atom, such as thienyl,etc.;

unsaturated 3- to 8-membered (more preferably 5- or 6-membered)heteromonocyclic group containing one oxygen atom, such as furyl, etc.;

unsaturated condensed heterocyclic group containing 1 or 2 sulfuratoms(s) and 1 to 3 nitrogen atom(s), such as benzothiazolyl,benzothiadiazolyl, etc.; and the like.

Suitable "acyl" may include lower alkanoyl such as formyl, acetyl,propionyl, butyryl, isobutyryl, valeryl, isovaleryl, pivaloyl, etc.;lower alkoxycarbonyl such as methoxycarbonyl, ethoxycarbonyl,propoxycarbonyl, isopropoxycarbonyl, butoxycarbonyl,tert-butoxycarbonyl, pentyloxycarbonyl, tert-pentyloxycarbonyl,hexyloxycarbonyl, etc.; lower alkylsulfonyl such as methylsulfonyl,ethylsulfonyl, propylsulfonyl, isopropyisulfonyl, butylsulfonyl,tert-butylsulfonyl, pentylsulfonyl, tert-pentylsulfonyl, hexylsulfonyl,etc.; arylsulfonyl such as phenylsulfonyl, naphthytsulfonyl, etc.; aroylsuch as benzoyl, naphthoyl, etc.; ar(lower)aikanoyl, such asphenylacetyl, phenylpropionyl, etc.; cyclo(lower)alkyl(lower)alkanoylsuch as cyclohexylacetyl, cyclopentylacetyl, etc.;ar(lower)alkoxycarbonyl such as benzyloxycarbonyl, phenethyloxycarbonyl,etc.; arylcarbamoyl such as phenylcarbamoyl, naphthylcarbamoyl, etc.;heterocyclicsulfonyl such as heteromonocyclic sulfonyl (e.g.thienyisulfonyl, furylsulfonyl, pyridylsulfonyl, etc.), etc.; and thelike.

The acyl group mentioned above may be substituted with 1 to 3 suitablesubstituent(s) such as halogen (e.g. chlorine, bromine, fluorine andiodine), lower alkyl (e.g. methyl, ethyl, propyl, isopropyl, butyl,isobutyl, sec-butyl, tert-butyl, pentyl, tert-pentyl, hexyl, etc.),lower alkoxy (e.g. methoxy, ethoxy, propoxy, isopropoxy, butoxy,tert-butoxy, pentyioxy, tert-pentyloxy, hexyloxy, etc.), nitro, mono-(ordi- or tri-)halo(lower)alkyl (e.g. chloromethyl, bromomethyl,chloropropyl, 1,2-dichloroethyl, 1,2-dibromoethyl, 2,2-dichloroethyl,trifluoromethyl, 1,2,2-trichloroethyl, etc.) or the like.

Suitable "protected carboxy" and "protected carboxy moiety" in the term"protected carboxy(lower)alkyl" may include carbamoyl; acylcarbamoylsuch as lower alkylsulfonylcarbamoyl (e.g. methylsulfonylcarbamoyl,ethylsulfonylcarbamoyl, propylsulfonylcarbamoyl,isopropylsulfonylcarbamoyl, butylsulfonylcarbamoyl,tert-pentylsulfonylcarbamoyl, pentylsulfonylcarbamoyl,tert-pentylsulfonylcarbamoyl, hexylsulfonylcarbamoyl, etc.),arylsulfonylcarbamoyl (e.g. phenylsulfonylcarbamoyl,naphthylsulfonylcarbamoyl, etc.) or the like; esterified carboxy inwhich said ester may be the ones such as lower alkyl ester (e.g. methylester, ethyl ester, propyl ester, isopropyl ester, butyl ester, isobutylester, tert-butyl ester, pentyl ester, tert-pentyl ester, hexyl ester,etc.), lower alkenyl ester (e.g. vinyl ester, allyl ester, etc.), loweralkynyl ester (e.g. ethynyl ester, propynyl ester, etc.), mono-(or di ortri)halo(lower)alkyl ester (e.g. 2-iodoethyi ester, 2,2,2-trichloroethylester, etc.), lower alkanoyloxy(lower)alkyl ester (e.g. acetoxymethylester, propionyloxymethyl ester, 1-acetoxypropyl ester, valeryloxymethylester, pivaloyloxymethyl ester, hexanoyloxymethyl ester, 1-acetoxyethylester, 2-propionyloxyethyl ester, 1-isobutyryloxyethyl ester, etc.),lower alkylsulfonyl(lower)alkyl ester (e.g. mesylmethyl ester,2-mesylethyl ester, etc.), ar(lower)alkyl ester such asphenyl(lower)alkyl ester which may be substituted by one or moresuitable substituent(s) (e.g. benzyl ester, 4-methoxybenzyl ester,4-nitrobenzyl ester, phenethyl ester, trityl ester, diphenylmethylester, bis(methoxyphenyl)methyl ester, 3,4-dimethoxybenzyl ester,4-hydroxy-3,5-di-tert-butylbenzyl ester, etc. or the like, loweralkoxycarbonyloxy(lower)aikyl ester (e.g. methoxycarbonyloxymethylester, ethoxycarbonyloxymethyl ester, ethoxycarbonyloxyethyl ester,etc.), aroyloxy(lower)alkyl ester (e.g. benzoyloxymethyl ester,benzoyloxyethyl ester, toluoyloxyethyl ester, etc.), aryl ester whichmay have one or more suitable substituent(s) (e.g. phenyl ester, tolylester, tert-butylphenyl ester, xylyl ester, mesityl ester, cumenylester, etc.), and the like.

Suitable "halogen" may include chlorine, bromine, fluorine and iodine.

Suitable "leaving group" may include acid residue and the like, and thesuitable examples thereof are halogen (e.g. chlorine, bromine, fluorine,etc.), sulfonyloxy (e.g. methylsulfonyloxy, phenylsulfonyloxy, etc.),and the like.

Suitable "anion" may include halide ion (e.g. chloride ion, bromide ion,fluoride ion, etc.), and the like.

Suitable "alkali metal" may include sodium, potassium, and the like.

Suitable pharmaceutically acceptable salts of the object compound (I)are conventional nontoxic salts and include a metal salt such as analkali metal salt (e.g. sodium salt, potassium salt, etc.) and alkalineearth metal salt (e.g. calcium salt, magnesium salt, etc.), ammoniumsalt an organic base salt (e.g. trimethylamine salt, triethylamine salt,pyridine salt, picoline salt, dicyclohexylamine salt,N,N'-dibenzylethylenediamine salt, etc.), an organic acid salt (e.g.acetate, maleate, tartrate, methanesulfonate, benzenesulfonate, formate,toluenesulfonate, trifluoroacetate, etc.), an inorganic acid salt (e.g.hydrochloride, hydrobromide, sulfate, phesphate, etc.), a salt with anamino acid (e.g. arginine, aspartic acid, glutamic acid, lysine, etc.),and the like.

The followings are the preferred examples of compound (I).

R¹ is unsaturated 5- or 6-membered heteromonocyctic(lower)alkylcontaining 1 to 4 nitrogen atom(s) more preferably pyridyl(lower)alkyl;most preferably pyridyl(C₁ -C₄)alkyl!;

R² is arylsulfonyl which may have 1 to 3 substituent(s) selected fromthe group consisting of halogen, lower alkyl, lower alkoxy and mono(ordi or tri)halo(lower)alkyl more preferably phenylsulfonyl which may have1 or 2 substituent(s) selected from the group consisting of halogen,lower alkyl, lower alkoxy and mono(or di or tri)halo(lower)alkyl; mostpreferably phenylsulfonyl which may have halogen!,

R³ is carboxy(iower)alkyl or protected carboxy(lower)alkyl morepreferably esterified carboxy(lower)alkyl; most preferably loweralkoxycarbonyl(lower)alkyl!.

The processes for preparing the object compound (I) and startingcompound (II) of the present invention are explained in detail in thefollowing.

Process (1)

The compound (I) or a salt themeof can be produced by reacting thecompound (II) or a salt thereof with the compound (III) or a saltthereof.

The reaction is generally carried out in the common solvent, such asacetone, dioxane, acetonitrile, chloroform, methylene chloride, ethylenechloride, tetrahydrofuran, ethyl acetate, N,N-dimethylformamide,dimethyl sulfoxide or any other solvent that does not interfere with thereaction.

While the reaction temperature is not critical, this reaction isgenerally conducted under cooling to warming.

This reaction is preferably conducted in the presence of an inorganicbase such as alkali metal hydroxide (e.g. sodium hydroxide, potassiumhydroxide, etc.), alkaline earth metal hydroxide (e.g. magnesiumhydroxide, potassium hydroxide, etc.), alkali metal carbonate (e.g.sodium carbonate, potassium carbonate, cesium carbonate, etc.), alkalineearth metal carbonate (e.g. magnesium carbonate, calcium carbonate,etc.), alkali metal hydrogencarbonate (e.g. sodiumhydrogencarbonate,potassium hydrogencarbonate, etc.), alkali metal acetate (e.g. sodiumacetate, potassium acetate, etc.), alkaline earth metal phosphate (e.g.magnesium phosphate, calcium phosphate, etc.), alkali metalhydrogenphosphate (e.g. disodium hydrogenphosphate, dipotassiumhydrogenphosphate, etc.), alkali metal hydride (e.g. sodium hydrideetc.), alkali metal (lower)alkoxide (e.g. sodium methoxide, sodiumethoxide, potassium methoxide, potassium ethoxide, potassium t-butoxide,etc.) and the like, or an organic base such as tri(lower)alkylamine(e.g. trimethylamine, triethylamine, diisopropylethylamine, etc.),pyridine, lutidine, picoline, dimethylaminopyridine,N-(lower)alkylmorpholine and the like (preferably in the presence of anorganic base).

Process (A)

The compound (Va) or a salt thereof can be produced by subjecting thecompound (IV) or a salt thereof to introduction reaction of the carboxyprotective group.

This reaction can be carried out by the procedure described inPreparation 1 or similar manners thereto. Process 1

The compound (VII) or a salt thereof can be produced by reacting thecompound (V) or a salt thereof with the compound (VI) or a salt thereof.

The reaction is generally conducted in the common solvent, such asacetone, dioxane, chloroform, methylene chloride, ethylene chloride,tetrahydrofuran, ethyl acetate, N,N-dimethylformamide, dimethylsulfoxideor any other solvent that does not interfere with the reaction.

While the reaction temperature is not critical, this reaction isgenerally carried out under cooling, at room temperature, under warmingor heating. Process (B)-2

The compound (IX) or a alt thereof can be produced by reacting thecompound (VII) or a salt thereof with the compound (VIII) or a saltthereof.

This reaction is generally conducted in the common solvent such asdichloromethane, chloroform, methylene chloride, ethylene chloride,tetrahydrofuran, N,N-dimethylformamide or any other solvent that doesnot interfere with the reaction.

While the reaction temperature is not critical this reaction isgenerally conducted under cooling or at room temperature.

This reaction is preferably conducted in the presence of an inorganicbase such as alkali metal hydroxide (e.g. sodium hydroxide, potassiumhydroxide, etc.), alkaline earth metal hydroxide (e.g. magnesiumhydroxide, calcium hydroxide, etc.), alkali metal carbonate (e.g. sodiumcarbonate, potassium carbonate, cesium carbonate, etc.), alkaline earthmetal carbonate (e.g. magnesium carbonate, calcium carbonate, etc.),alkali metal hydrogencarbonate (e.g. sodium hydrogencarbonate, potassiumhydrogencarbonate, etc.), alkali metal acetate (e.g. sodium acetate,potassium acetate, etc.), alkaline earth metal phosphate (e.g. magnesiumphosphate, calcium phosphate, etc.), alkali metal hydrogenphosphate(e.g. disodium hydrogenphosphate, dipotassium hydrogenphosphate, etc.)and the like, or an organic base such as trialkylamine (e.g.trimethylamine, triethylamine etc.) and the like.

Process (B)-3

The compound (X) or a salt thereof can be produced by subjecting thecompound (IX) or a salt thereof to halogenation reaction.

This reaction can be carried out by the procedure described inPreparation 3 or similar manners thereto.

It should be noted that this reaction reverses the configulation of thesubstituent in 4-position of the pyrrolidine ring.

Process (B)-4

The compound (XII) or a salt thereof can be produced by reacting thecompound (X) or a salt thereof with the compound (XI).

This reaction is generally carried out in the common solvent such asdimethyl sulfoxide or any other solvent that does not interfere with thereaction.

The reaction temperature is not critical but the reaction is generallycarried out under warming or under heating. This reaction reverses theconfigulation of the substituent in 4-position of the pyrrolidine ring.

Process (B)-5

The compound (XIII) or a salt thereof can be produced by subjecting thecompound (XII) or a salt thereof to hydrogenation reaction.

This reaction can be conducted by the procedure described in Preparation5 or similar manners thereto.

Process (B)-6

The compound (XIV) or a salt thereof can be produced by reacting thecompound (XIII) or a salt thereof with an acylating agent.

The acylating agent is an organic acid, i.e. R² -OH (wherein R² is anacyl group), or its reactive derivative or a salt thereof. The preferredexamples of said reactive derivative of organic acid includes thederivatives commonly employed, such as the acid halide (e.g. acidchloride, acid bromide, etc.), acid azide, acid anhydride, activatedamide, activated ester, isocyanate, for example, an aryl isocyanate(e.g. phenyl isocyanate, etc.), and the like.

When the free acid is used as the acylating agent, the acylationreaction is preferably conducted in the presence of the conventionalcondensing agent such as N,N'-dicyclohexylcarbodiimide, etc.

The reaction is preferably carried out in the presence of an inorganicor organic base such as those mentioned for process (B)-2.

This reaction is generally conducted in a solvent which does notinterfere with the reaction, such as water, methanol, ethanol, propanol,dichloromethane, tetrahydrofuran, chloroform, ethyl acetate and thelike.

The reaction temperature is not critical and the reaction can be carriedout under cooling to heating.

Process (B)-7

The compound (XII) or a salt thereof can be produced by reacting thecompound (IX) or a salt thereof with the compound (XI).

This reaction can be carried out in the same manner to that of ProcessB-4 and, therefore, the reaction conditions can be referred to those ofthe Process (B)-4.

It should be noted that this reaction reverses the configulation of thesubstituent in 4-position of the pyrrolidine ring.

Process (C)

The compound (II) or a salt thereof can be produced by subjecting thecompound (XIV) or a salt thereof to reduction reaction.

This reduction reaction is generally carried out using a reducing agentsuch as di(lower)alkylaluminum hydride (e.g. diisobutylaluminum hydride,etc.), an alkali metal aluminum hydride (e.g. lithium aluminum hydride,sodium aluminum hydride, potassium aluminum hydride etc.), and the like.

This reaction is generally conducted in the common solvent such asmethylene chloride, chloroform, toluene, tetrahydrofuran, or any othersolvent that does not interfere with the reaction.

The reaction temperature is not critical and the reaction is generallycarried out under cooling or at room temperature.

The object compound (I) of this invention and pharmaceuticallyacceptable salts thereof are thromboxane A₂ (TXA₂) antagonists and TXA₂synthetase inhibitors and, therefore, they are useful for theprophylaxis and/or therapy of thrombotic diseases (for example,transient cerebral ischemic attack, cerebral apoplexy, unstable angina,myocardial infarction, peripheral circulatory insufficiency, thrombusformation after percutaneous transluminal coronary angioplasty,disseminated intravascular coagulation syndrome, etc.), allergicdiseases (e.g. asthma, etc.), nephritis, peptic ulcer, hemicrania,diabetic neuropathy, diabetic angiopathy, restenosis after percutaneoustransluminal coronary angioplasty, adult respiratory distress syndrome,shock, hepatic disorders (e.g. hepatitis, etc.), cerebral vasospasmafter subarachnoidal hemorrhage, hypertension, arteriosclerosis,cancerous metastasis, thrombus formation on extracorporeal circulation,thrombus formation on transplantation, conjunctivitis, etc. and forreducing nephrotoxicity induced by immunosuppressant drugs such ascyclosporin at renal transplantation, and can be also used withfibrinolytic agents in order to increase the effect of fibrinolyticagents.

Furthermore, compound (I) and pharmaceutically acceptable salts thereofare useful for the prophylaxis and/or therapy of cerebral infarctionsuch as acute cerebral infarction, arrhythmia, angina pectoris and so

The following test examples indicate that Compound (I) andpharmaceutically acceptable salts thereof are useful for the prophylaxisand/or therapy of cerebral infarction, arrhythmia, angina pectoris andso on.

Test Compound

(2S,4R)-2-(Z)-5-Carboxy-1-pentenyl!-4-(4-chloro-phenylsulfonylamino)-1-(3-pyridylmethyl)pyrrolidinehydrochloride (hereinafter referred to briefly as test compound (I)).

Test Example 1 Effect on the Cerebral Infarct Following Ligation ofMiddle Cerebral Artery in SHR rats

Nine-week-old male spontaneously hypertensive rats (SHRs, Charles RiverJapan Inc.) were used in the test.

After induction of anesthesia with 4% halothane (100% O₂) in rats, themiddle cerebral artery was occluded with an electric coagulator.Immediately after occlusion, the introduction of halothane was stoppedand the incised skin area was sutured. Test compound (I) was suspendedin 0.5% methylcellulose solution and the suspension was administeredorally in a dosing volume of 5 ml/kg immediately after artery occlusion.

Twenty-four hours after occlusion of the middle cerebral artery, thebrain was removed from the rat and after the bregma was located, acoronary section 2 mm posterior to the bregma was prepared. This sectionwas immersed in 2% triphenyltetrazolium chloride (TTC) solution andmaintained at 37° C. for 40 minutes. After staining, the cerebralsection was photographed and the area ratio of the infarct to thehemisphere (%) was calculated for each cut surface using a computerizedanalyzer.

For statistical analysis, the Mann-Whitney U-test was used.

Results

    ______________________________________                                        Dose        Number of                                                         (mg/kg)     animals   Item                                                    ______________________________________                                        Control                                                                               0       7         Non-infarcted                                                                            43.167 ±                              group                     area (mm.sup.2)                                                                          1.253                                                              Area of cortical                                                                         12.116 ±                                                        infarct (mm.sup.2)                                                                       1.302                                                              Total area 55.283 ±                                                        (mm.sup.2) 1.217                                                              % Infarct  21.823 ±                                                                   2.194                                    Drug-  320      9         Non-infarcted                                                                            **49.299 ±                            treated                   area (mm.sup.2)                                                                          1.301                                    group                     Area of cortical                                                                         9.103 ±                                                         infarct (mm.sup.2)                                                                       0.491                                                              Total area 58.402 ±                                                        (mm.sup.2) 1.203                                                              % Infarct  *15,643 ±                                                                  0.892                                    ______________________________________                                         Mean ± SE                                                                  **p < 0.01 compared with control                                              *p < 0.05 compared with control                                          

Test Example 2 Effect on Arrhythmia Following CoronaryIschemia-Reperfusion in Rats

Male Wistar rats (11 weeks old) fasted for 24 hours were anesthetizedwith pentobarbital Na, 50 mg/kg i.p. After thoracotomy under artificialrespiration, the descending branch of the coronary artery was compressedby suction to arrest the blood flow. After 5 minutes perfusion wasreestablished and the animals were observed for ventricular fibrillation(VF) and death for 5 minutes.

Test compound (I) was administered orally 1 hour before arrest of bloodflow.

Results

    ______________________________________                                                 Dose   Number of Incidence Mortality                                          (mg/kg)                                                                              animals   of VF (%) rate (%)                                  ______________________________________                                        Control group                                                                            --       17        94.1    52.9                                    Drug-treated group                                                                       100       6        50*     0*                                      ______________________________________                                         *p < 0.05 compared with control                                          

Test Example 3 Effect on Vasopressin-Induced Angina in Angina Model Rats

Mole Donryu rats (6 weeks old) fasted for 24 hours were anesthetizedwith pentobarbital Na, 60 mg/kg i.p. Then, each animal was fixed insupine position and needle electrodes for ECG were affixed to the fourlimbs. ECG was recorded in Lead II. Vasopressin (Sigma), 0.21 U/kg, wasadministered into the femoral vein and ECG changes were recorded.

The onset of angina was assessed by a depression of ST segment (ΔST(ΔV)).

Test compound (I) was administered orally 1 hour before vasopressinadministration.

    ______________________________________                                                   Dose      Number of                                                                              Δ ST                                                 (mg/kg)   animals  (μV)                                         ______________________________________                                        Control group                                                                              --          7        136 ± 13                                 Drug-treated group                                                                         320         8         62 ± 26                                 ______________________________________                                         Mean ± SE                                                             

The object compound (I) or a pharmaceutically acceptable salt thereofcan usually be administered to mammals including human being, generallyin the form of a conventional pharmaceutical composition such ascapsule, micro-capsule, tablet, granule, powder, troche, syrup, aerosol,inhalation, solution, injection, eye-drop, nasal drop, suspension,emulsion, suppository, ointment, or the like.

The pharmaceutical composition of this invention may contain a variousorganic and/or inorganic carrier substances which are commonly used inpharmaceutical preparations such as excipient (e.g. sucrose, starch,mannit, sorbit, lactose, glucose, cellulose, talc, calcium phosphate,calcium carbonate, etc.), binding agent (e.g. cellulose,methylcellulose, hydroxypropylcellulose, polypropylpyrrolidone, gelatin,gum arabic, polyethylene glycol, sucrose, starch, etc.), disintegratingagent (e.g. starch, carboxymethylcellulose, calciumcarboxymethylcellulose, hydroxypropylstarch, glycol-starch sodium,sodium hydrogencarbonate, calcium phosphate, calcium citrate, etc.),lubricant (e.g. magnesium stearate, talc, sodium lauryl sulfate, etc.),flavoring agent (e.g. citric acid, menthol, glycine, orange powder,etc.), preservative (e.g. sodium benzoate, sodium hydrogen sulfite,methytparaben, propylparaben, etc.), stabilizer (e.g. citric acid,sodium citrate, acetic acid, etc.), suspending agent (e.g.methylcellulose, polyvinylpyrrolidone, aluminum stearate, etc.),dispersing agent, aqueous diluting agent (e.g. water, etc.), base wax(e.g. cacao butter, polyethylene glycol, white petrolatum, etc.) and thelike.

The effective ingredient may usually be administered with a unit dose of0.01 mg/kg to 50 mg/kg, 1 to 4 times a day. However, the above dosagemay be increased or decreased according to age, weight, conditions ofthe patient or the administering method.

The following preparations and examples are given for the purpose ofillustrating the present invention in more detail.

Preparation 1

In an atmosphere of nitrogen, (2S,4R)-2-carboxy-4-hydroxypyrrolidine(100 g, 0.763 mol) was suspended in methanol (400 ml) and the suspensionwas cooled to 5°-10° C. Then, thionyl chloride (99.8 g, 0.839 mol) wasadded dropwise thereto under 20° C. After completion of the dropwiseaddition, the reaction mixture was warmed to 60°-62° C. and stirred atthis temperature for 1 hour. The reaction mixture was then cooled to25°-30° C. for crystallization. To the mixture was added diisopropylether (180 ml) dropwise. After completion of the dropwise addition, themixture was cooled to 0°-5° C. and stirred for 1 hour. The resultingcrystal was collected by filtration and washed with diisopropyl ether(200 ml) twice. The washed crystal was dried in vacuo overnight toprovide (2S,4R)-2-methoxycarbonyl-4-hydroxypyrrolidine hydrochloride(134.8 g).

IR (Nujol): 3320, 1740, 1590, 1080, 1620, 900 cm⁻¹

NMR (DMSO-d₆, δ): 5.84 (1H, s), 4.50-4.40 (1H, m), 4.42 (1H, s), 3.70(1H, d, J=12 Hz), 3.60 (1H, d,

J=12 Hz), 3.07 (1H, d, J=12 Hz), 2.22-2.20 (1H, m)

Preparation 2

In an atmosphere of nitrogen, 3-hydroxymethylpyridine (35.95 g, 0.329mol) was dissolved in N,N-dimethylformamide (250 ml) followed by coolingto -15°˜20° C. Then, triethylamine (36.65 g, 0.329 mol) was addedthereto and methanesulfonyl chloride (41.45 g, 0.362 mol) was addeddropwise to the mixture at 0°˜-10° C. After completion of the dropwiseaddition, the mixture was stirred at -10° C. for 30 minutes. To thismixture were added (2S,4R)-2-methoxycarbonyl-4-hydroxypyrrolidinehydrochloride (50 g, 0.275 mol) and triethylamine (100 g, 0.98.8 mol)followed by warming to 65° C. The mixture was stirred at 65° C. for 1hour to provide a reaction mixture containing(2S,4R)-1-(3-pyridylmethyl)-2-methoxycarbonyl-4-hydroxypyrrolidine. Thisreaction mixture was cooled to -20° C. and triethylamine (66.8 g, 0.66mol) was added thereto. Then, methanesulfonyl chloride (75.6 gt 0.66mol) was added thereto dropwise at -20°˜15° C. The mixture was stirredat -20°˜25° C. for 1 hour, after which it was poured into ethyl acetate(1 l)-water (1 l ) and extracted. After phase separation, the aqueouslayer was extracted with ethyl acetate (500 ml) twice. The ethyl acetatelayer was combined and washed with a saturated aqueous sodium chloridesolution (125 ml). The ethyl acetate layer was concentrated to drynessunder reduced pressure to provide(2S,4R)-1-(3-pyridylmethyl)-2-methoxycarbonyl-4-methylsulfonyloxypyrrolidine(84.65 g) as an oil.

NMR (CD₃ OD, δ): 9.8-9.6 (2H, m), 8.2-8.0 (1H, m), 7.6 (1H, m), 5.25(1H, q, J=3.6Hz), 4.19, 4.08, 3.86, 3.75 (2H, ABq), 3.68 (3H, s), 3.42(1H, dd, J=5.8 Hz, J=12Hz), 3.05 (3H, s), 3.00 (1H, s), 2.40-2.55 (2H,m)

Preparation 3

In an atmosphere of nitrogen,(2S,4R)-1-(3-pyridylmethyl)-2-methoxycarbonyl-4-methylsulfonyloxypyrrolidine(84.65 g) was dissolved in polyethylene glycol-400 (400 ml) followed byaddition of lithium chloride (40 g, 0.943 mol). The mixture was warmedto 85°-90° C. and stirred at that temperature for 4-5 hours. Thereaction mixture was then cooled to -20°˜-25° C. and ethyl acetate (800ml)-water (400 ml) was added thereto for extraction. After phaseseparation, the aqueous layer was adjusted to pH 9-9.5 with 24% sodiumhydroxide in water (pH prior to adjustment: 4.8) and extracted withethyl acetate (400 ml) twice. The ethyl acetate layer was combined andwashed with a saturated aqueous sodium chloride solution (200 ml). Theethyl acetate solution was then concentrated to dryness under reducedpressure to provide(2S,4S)-1-(3-pyridylmethyl)-2-methoxycarbonyl-4-chloropyrrolidine (60.6g) as an oil.

NMR (CDCl₃, δ): 8.55 (2H, m), 7.50 (1H, m), 7.30 (1H, m), 4.20 (1H, m),4.00-4.15 (2H, m), 3.73 (3H, s), 3.75 (1H, m), 3.30 (1H, m), 3.05 (1H,m), 2.95 (1H, m), 2.20 (1H, m)

Preparation 4

In an atmosphere ofnitrogen,(2S,4S)-1-(3-pyridylmethyl)-2-methoxycarbonyl-4-chloropyrrolidine(60.6 g) was dissolved in dimethyl sulfoxide (600 ml) followed byaddition of sodium azide (60 g, 0,923 mol) and the mixture was warmed to85°-90° C. and stirred at the same temperature for 5-6 hours. Thereaction mixture was cooled to 40°-50° C. and poured into ethyl acetate(1 l)-water (1 l) for extraction. After phase separation, the aqueouslayer was re-extracted with ethyl acetate (500 ml). The ethyl acetatelayer was combined and washed with a saturated aqueous sodium chloridesolution (250 ml). The ethyl acetate solution was concentrated todryness under reduced pressure to provide(2S,4R)-1-(3-pyridylmethyl)-2-methoxycarbonyl-4-azidopyrrolidine (49.4g) as an oil.

NMR (CDCl₃, δ): 8.55 (2H, m), 7.70 (1H, m), 7.26 (1H, m), 4.10 (1H, m),3.97, 3.90, 3.71, 3.64 (2H, ABq), 3.68 (3H, s), 3.35 (1H, m), 2.60 (1H,m), 2.00-2.40 (2H, m)

Preparation 5

In an atmosphere ofnitrogen,(2S,4R)-1-(3-pyridylmethyl)-2-methoxycarbonyl-4-azidopyrrolidine(49.4 g) was dissolved in ethyl acetate (500 ml) followed by addition oftriphenylphosphine (60.75 g, 0.232 mol) at 20°-30° C. (foaming tookplace). After completion of addition, the mixture was warmed to 40°-45°C. and stirred for 30 minutes. The mixture was diluted with water (16.5ml) and warmed and stirred at 60°-65° C. for 1.5 hours to provide areaction mixture containing(2S,4R)-1-(3-pyridylmethyl)-2-methoxycarbonyl-4-aminopyrrolidine. Thisreaction mixture was cooled to 5°-10° C. and adjusted to pH 3.5 with 1Nhydrochloric acid (ca. 350 ml). After phase separation, the organiclayer was washed with 1% hydrochloric acid (125 ml). The aqueous layerwas combined and adjusted to pH 7.0 with triethylamine (ca. 20 ml) (pHprior to adjustment: 1.5). This aqueous solution was cooled to 5° C. andethyl acetate (475 ml) and triethylamine (70.5 g, 0.696 mol) were addedthereto in that order. Then, at 5°-10° C., 4-chlorobenzenesulfonylchloride (49 g, 0.232 mol) was added in 3 successive portions and themixture was stirred at 5° C. for 1 hour. The reaction mixture wasadjusted to pH 2.0 with 6N-hydrochloric acid. After phase separation,the organic layer was washed with 1% hydrochloric acid (125 ml). Theaqueous layer was combined and adjusted to pH 7.0 with 24% aqueoussodium hydroxide solution. The aqueous solution was extracted with ethylacetate (500 ml) and the extract was washed with a saturated aqueoussodium chloride solution (125 ml). The ethyl acetate layer wasconcentrated to 187.5 ml under reduced pressure and the concentrate wasstirred at 20°-25° C. for 1.5 hours for crystallization and ripening. Tothis solution was added diisopropyl ether (562.5 ml) dropwise and themixture was stirred for 1 hour. The resulting crystal was collected byfiltration and washed with diisopropyl ether (100 ml). The washedcrystal was dried in vacuo overnight to provide(2S,4R)-1-(3-pyridylmethyl)-2-methoxycarbonyl-4-(4-chlorophenyl-sulfonylamino)pyrrolidine(51.51 g) as a crystal.

IR (Nujol) : 1740, 1590, 1340, 1160 cm⁻¹

NMR (CDCl₃, δ): 8.51-8.45 (2H, m), 7.70 (2H, d, J=11.3Hz), 7.63-7.58(1H, m), 7.42 (2H, d, J=11.3Hz), 7.28-7.21 (1H, m), 6.30 (1H, d, J=8Hz), 3.95 (1H, m), 3.85, 3.78, 3.65, 3.58 (2H, ABq), 3.65 (3H, s), 3.50(1H, m), 3.15 (1H, m), 2.40-2.20 (2H, m), 2.00 (1H, m)

Preparation 6

In an atmosphere of nitrogen,(2S,4R)-1-(3-pyridylmethyl)-2-methoxycarbonyl-4-(4-chlorophenylsulfonylamio)pyrrolidine(10 g, 0.0244 mol) was dissolved in methylene chloride (200 ml) and thesolution was cooled to -50°˜-55° C. Then, 1M diisobutylaluminum hydridein toluene (73.2 ml, 0.0732 mol) was added thereto dropwise at -50° C.and the mixture was stirred for 30 minutes. Then, 20% sodium potassiumtartrate solution was added thereto at 25° C. for precipitation ofinsolubles. The insolubies were separated by filtration and washed withmethylene chloride. The mother liquor and the washings were combinedand, after phase separation, the methylene chloride layer was taken. Themethylene chloride layer was concentrated to dryness to provide(2S,4R)-1-(3-pyridylmethyl)-2-formyl-4-(4-chlorophenylsulfonylamino)-pyrrolidine(10.4 g) as an oil.

NMR (CDCl₃, δ): 9.37 (1H, d, J=2.6Hz), 8.50 (2H, m), 7.75 (2H, d, J=11.3Hz), 7.60 (1H, m), 7.20-7.10 (1H, m), 7.40 (2H, d, J=11.3 Hz), 3.81,3.74, 3.65, 3.58 (2H, ABq), 3.25 (1H, m), 2.20 (1H, m), 1.90 (1H, m)

EXAMPLE 1

In an atmosphere of nitrogen, potassium t-butoxide (21.9 g, 0.195 mol)was dissolved in tetrahydrofuran (140 ml) and after the solution wascooled to 0°-5° C., (4-carboxybutyl)triphenylphosphonium chloride (43.3g, 0.0975 mol) was added thereto. The mixture was warmed to 40° C. andstirred for 1 hour. The reaction mixture was cooled to 0°-5° C. and asolution of(2S,4R)-1-(3-pyridylmethyl)-2-formyl-4-(4-chlorophenylsulfonylamino)pyrrolidine(10.4 g) in tetrahydrofuran (60 ml) was added thereto dropwise. Thisreaction mixture was stirred at -5°˜5° C. for 1 hour, after which it waspoured into methylene chloride (200 ml) and 1N-hydrochloric acid (400ml) for extraction. After phase separation, the methylene chloride layerwas extracted with 1N-hydrochloric acid (200 ml) twice. The aqueouslayer was combined and adjusted to pH 5.5 with 24% sodium hydroxide inwater (pH prior to adjustment=0.25-0.3). The aqueous solution wasextracted with ethyl acetate (200 ml), while the aqueous layer wasre-extracted with ethyl acetate (100 ml). The ethyl acetate layer wascombined and concentrated to dryness under reduced pressure to providean oil. This oil was dissolved in 1N-hydrochloric acid (70 ml)-purifiedwater (35 ml) and the solution was adjusted to pH 2.2 by dropwiseaddition of 1N-aqueous sodium hydroxide solution. The mixture wasstirred at 20°-25° C. for crystallization and ripening. During thisprocedure, 1N-aqueous sodium hydroxide solution was added theretodropwise so as to maintain the solution at pH 2.2. After (about 1 hourof) ripening, 1N-aqueous sodium hydroxide solution was added theretodropwise so as to adjust the solution to pH 3.5. The solution was thencooled to 5° C. and stirred for 1 hour. The resulting crystal wascollected by filtration, washed with cold purified water (10 ml) anddried in vacuo overnight to provide (2S,4R)-2-(Z)-5-carboxy-1-pentenyl!-4-(4-chlorophenylsulfonylamino)-1-(3-pyridylmethyl)pyrolidinehydrochloride (8.0 g) as crude crystal.

IR (Nujol): 3100, 1700, 1340, 1180, 1160, 1000 cm⁻¹

NMR (DMSO-d₆, δ): 12.0 (1H, br s), 8.85 (3H, m), 8.05 (1H, m), 7.85 (2H,d, J=8.8 Hz), 7.70 (2H, d, J=8.8 Hz), 7.50 (1H, m), 5.70 (2H, m), 4.60(1H, m), 4.30 (2H, m), 3.95 (1H, m), 3.35 (1H, m), 2.90 (1H, m),2.30-1.80 (6H, m), 1.60 (2H, m)

Example 2

In 1N-hydrochloric acid (44 ml)-purified water (16 ml) was dissolvedcrude crystal of (2S,4R)-2-(Z)-5-carboxy-1-pentenyl!-4-(4-chlorophenylsulfonylamino)-1-(3-pyridylmethyl)pyrrolidinehydrochloride (8.0 g, 0.016 mol) followed by addition of activatedcarbon (0.8 g) and the mixture was stirred at 25° C. for 1 hour. Then,clarification-filtration was carried out using a 0.25μmembrane prefilterand the filtrate was washed with 1N-hydrochloric acid (4 ml)-purifiedwater (8 ml). While the filtrate was stirred, 1N-aqueous sodiumhydroxide solution was added dropwise so as to adjust the solution to pH2.0 (pH prior to adjustment: 0.4). The mixture was stirred at 20°-25° C.for crystallization and ripening (about 1 hour, pH decreases to 1.51).Then, 1N-aqueous sodium hydroxide solution was added thereto dropwise soas to adjust the solution to pH 2.20. The solution was cooled to 5° C.and 1N aqueous sodium hydroxide solution was added thereto dropwise soas to maintain the solution at pH 2.20. The solution was cooled to 5° C.and 1N-aqueous sodium hydroxide solution was added thereto dropwise soas to maintain the solution at pH 2.20. The solution was then allowed tostand at 0°-5° C. overnight. The next morning the crystal was collectedby filtration and washed with cold purified water (8 ml). The washedcrystal was dried in vacuo overnight to provide pure crystal (6.77 g) of(Z)-5-carboxy-l-pentenyl!-4-(4-chlorophenylsulfonylamino)-1-(3-pyridylmethyl)pyrrolidinehydrochloride.

We claim:
 1. A compound of the general formula: ##STR5## wherein R¹ ispyridyl(lower)alkyl; and R² is acyl;or a salt thereof.
 2. A compound ofclaim 1,wherein R¹ is pyridyl(lower)alkyl; and is phenylsulfonyl whichmay have halogen.
 3. A compound of claim 2, wherein R¹ ispyridyl(lower)alkyl; andR² is halophenylsulfonyl.
 4. A compound of claim2, which is(2S,4R)-1-(3-pyridylmethyl)-2-formyl-4-(4-chlorophenylsulfonylamino)pyrrolidine.5. A process for producing a compound of the general formula: ##STR6##wherein R¹ is pyridyl(lower)alkyl; and R² is acyl;or a salt thereofcharacterized by subjecting a compound of the general formula: ##STR7##wherein R¹ is pyridyl(lower)alkyl; R² is acyl; and R⁵ is carboxy orprotected carboxy;or a salt thereof to reduction reaction using areducing agent which is a di(lower)alkyl aluminum hydride or a alkalimetal aluminum hydride.
 6. The process of claim 5, wherein said reducingagent is diisobutyl aluminum hydride, lithium; aluminum hydride, sodiumaluminum hydride, or potassium aluminum hydride.